Rexinoid bexarotene modulates triglyceride but not cholesterol metabolism via gene-specific permissivity of the RXR/LXR heterodimer in the liver

Abstract

Objective: Bexarotene (Targretin) is a clinically used antitumoral agent which exerts its action through binding to and activation of the retinoid-X-receptor (RXR). The most frequent side-effect of bexarotene administration is an increase in plasma triglycerides, an independent risk factor of cardiovascular disease. The molecular mechanism behind this hypertriglyceridemia remains poorly understood.

Methods and results: Using wild-type and LXR alpha/beta-deficient mice, we show here that bexarotene induces hypertriglyceridemia and activates hepatic LXR-target genes of lipogenesis in an LXR-dependent manner, hence exerting a permissive effect on RXR/LXR heterodimers. Interestingly, RNA analysis and Chromatin Immunoprecipitation assays performed in the liver reveal that the in vivo permissive effect of bexarotene on the RXR/LXR heterodimer is restricted to lipogenic genes without modulation of genes controlling cholesterol homeostasis.

Conclusions: These findings demonstrate that the hypertriglyceridemic action of bexarotene occurs via the RXR/LXR heterodimer and show that RXR heterodimers can act with a selective permissivity on target genes of specific metabolic pathways in the liver.

Figure 1. RXR or LXR agonist treatment increases plasma and hepatic triglyceride levels in vivo in an LXRα/β-dependent manner

Plasma (A) and hepatic (B) triglyceride concentrations were measured in LXRα/β-deficient mice (□) and wild type mice (■) gavaged with the RXR agonist (bexarotene, 30 and 100 mpk), the LXR agonist (T0901317, 30 mpk) or vehicle alone for 14 days (N=7/group). **p<0.01, ***p<0.001 versus wild type mice treated with vehicle; ^§p<0.05, ^§§p<0.01 versus LXRα/β-deficient mice treated with vehicle.

Figure 2. RXR or LXR agonist treatment induces hepatic lipogenic gene expression in mice in vivo in an LXRα/β-dependent manner

Hepatic mRNA levels of SREBP1c, SCD1 and FAS were measured in LXRα/β-deficient mice (□) and wild type mice (■) gavaged with the RXR agonist (bexarotene, 30 and 100 mpk), the LXR agonist (T0901317, 30 mpk) or vehicle alone for 14 days (N=7/group). Data normalized to 28S mRNA are expressed as a percentage of the wild type mice group treated with vehicle alone, to which an arbitrary value of 100 was attributed. **p<0.01 versus wild type mice treated with vehicle, ^§p<0.05, ^§§p<0.01 versus LXRα/β-deficient mice treated with vehicle.

Figure 3. RXR or LXR agonist treatment increases RXR recruitment to the LXREs of lipogenic genes in vivo

ChIP assays were performed using antibodies against RXR (A) or RNA polymerase II (B) on livers of LXRα/β-deficient mice and wild type mice treated with vehicle alone, bexarotene (100 mpk) or T0901317 (30 mpk). Recovery was determined by real-time PCR using primers positioned at the LXRE of SREBP1c, FAS and SCD1 (A) or primers located in the body of the gene (B). β-globin was used as a control. *p<0.05, **p<0.01, ***p<0.001 versus wild type mice treated with vehicle, ^§p<0.05, ^§§p<0.01, ^§§§p<0.001 versus LXRα/β-deficient mice treated with vehicle.

Figure 3. RXR or LXR agonist treatment increases RXR recruitment to the LXREs of lipogenic genes in vivo

ChIP assays were performed using antibodies against RXR (A) or RNA polymerase II (B) on livers of LXRα/β-deficient mice and wild type mice treated with vehicle alone, bexarotene (100 mpk) or T0901317 (30 mpk). Recovery was determined by real-time PCR using primers positioned at the LXRE of SREBP1c, FAS and SCD1 (A) or primers located in the body of the gene (B). β-globin was used as a control. *p<0.05, **p<0.01, ***p<0.001 versus wild type mice treated with vehicle, ^§p<0.05, ^§§p<0.01, ^§§§p<0.001 versus LXRα/β-deficient mice treated with vehicle.

Figure 4. LXR, but not RXR agonist treatment increases plasma cholesterol levels in vivo

TC (A), HDL-C (B) and non-HDL-C (C) were measured in LXRα/β-deficient mice (□) and wild type mice (■) gavaged, as indicated, with the RXR agonist (bexarotene, 30 and 100 mpk), the LXR agonist (T0901317, 30 mpk) or vehicle alone for 14 days (N=7/group). **p<0.01, ***p<0.001 versus wild type mice treated with vehicle.

Figure 5. LXR, but not RXR agonist treatment upregulates hepatic cholesterol genes in vivo

Hepatic mRNA levels of ABCG1, ABCA1, ABCG5 and ABCG8 were measured in LXRα/β-deficient mice (□) and wild type mice (■) gavaged, as indicated, with the RXR agonist (bexarotene, 30 and 100 mpk), the LXR agonist (T0901317, 30 mpk) or vehicle alone for 14 days (N=7/group). Data normalized to 28S gene are expressed as a percentage of the wild type mice group treated with vehicle alone, to which an arbitrary value of 100 was attributed. * p<0.05, **p<0.01 versus wild type mice treated with vehicle, ^§p<0.05 versus LXRα/β-deficient mice treated with vehicle.

Figure 6. LXR, but not RXR agonist treatment increases RXR recruitment to the LXRE of cholesterol genes in vivo

ChIP assays were performed using antibodies against RXR (A) or RNA polymerase II (B) on livers of LXRα/β-deficient mice and wild type mice treated with vehicle alone, bexarotene (100 mpk) or T0901317 (30 mpk). Recovery was determined by real-time PCR using primers positioned at the LXRE of ABCA1, ABCG1 and at two putative LXREs of ABCG5 and ABCG8 (A) or primers located in the body of the gene (B). β-globin was used as a control. *p<0.05, **p<0.01 versus wild type mice treated with vehicle.

Figure 6. LXR, but not RXR agonist treatment increases RXR recruitment to the LXRE of cholesterol genes in vivo

ChIP assays were performed using antibodies against RXR (A) or RNA polymerase II (B) on livers of LXRα/β-deficient mice and wild type mice treated with vehicle alone, bexarotene (100 mpk) or T0901317 (30 mpk). Recovery was determined by real-time PCR using primers positioned at the LXRE of ABCA1, ABCG1 and at two putative LXREs of ABCG5 and ABCG8 (A) or primers located in the body of the gene (B). β-globin was used as a control. *p<0.05, **p<0.01 versus wild type mice treated with vehicle.